Transcript Document
Chapter 16 The Genetic Basis of Development zygote adult 4 and 6 April, 2005 Overview • Instructions in the genome establish the developmental fate of cells in multicellular organisms. • Developmental pathways consist of sequences of various regulatory steps. • The zygote is totipotent, giving rise to all body cells. • Gradients of maternally-derived regulatory proteins establish polarity of the body axis and control transcriptional activation of zygotic genes. • Transcriptional regulation and cell signaling mediate development in animals and plants. • The same set of genes appears to regulate early development in all animals. Development • In multicellular organisms, life begins as a single cell. • With few exceptions, somatic cells contain the same genetic information as the zygote. • In development, cells commit to specific fates and differentially express subsets of genes. • Cells identify and respond to their position in developmental fields. • Daughter cells may differ with respect to regulatory instructions and developmental fate. Building the embryo • Developmental decisions – made at specific times during development – many are binary, e.g., male or female, germ line or somatic. – most are irreversible – many involve groups of cells rather than single cells • In animals decisions are made to – establish anterior-posterior and dorsal-ventral axes – subdivide anterior-posterior axis into segments – subdivide dorsal-ventral axis into germ layers – produce various tissues and organs • Most decisions involve changes in transcription Sex determination • XX-XY chromosomal systems for sex determination have evolved many times • Different molecular pathways for sex determination in different groups of animals • Drosophila – each cell lineage makes sexual decision – ratio of X chromosomes to autosomes determines sex – cascade of differential mRNA splicing • Mammals – TDF gene on Y chromosome determines maleness – endocrine hormonal system Sxl toggle • Ratio of NUM bHLH proteins to DEM bHLH proteins measures X:A ratio by competing for dimer formation – DNA binding domain of NUM proteins recognizes Sxl early promoter – twice as much NUM protein in females with two X chromosomes as males with one X results in more NUMNUM homodimers • Sufficient NUM-NUM homodimers activate Sxl early promoter resulting in SXL protein that alternatively splices larger Sxl transcript from late promoter – sets up autoregulatory loop in flies with X:A ratio of 1.0 – in flies with X:A ratio of 0.5, insufficient NUM-NUM homodimers results in no SXL protein and late transcript is normally processed (yields nonfunctional protein) Sxl downstream target • SXL protein activates downstream shunt that leads to female development – SXL protein binds to primary transcript of tra (transformer) resulting in spliced transcript that produces TRA protein – TRA protein in turn is RNA-binding protein that produces femalespecific splicing of dsx (doublesex) transcript – DSX-F transcription factor represses male-specific gene expression resulting in female development • In absence of SXL, there is no functional TRA protein, and dsx is spliced to produce DSX-M transcription factor which represses female-specific genes, leading to male development Sex determination in mammals • Presence of Y chromosome determines maleness – SRY gene in humans encodes transcription factor (testis-determining factor) – expression of SRY in developing gonad causes it to develop into testis – testis secretes testosterone resulting in male development • In XX individuals, absence of SRY protein and subsequent absence of testosterone results in default female shunt pathway Role of cytoskeleton in development • Consists of highly organized rods and fibers – microfilaments (actin) – intermediate filaments – microtubules • Such structures are polar, with distinct “+” and “–” ends • Serve as highway system for intracellular transport • Asymmetry of cytoskeletal elements plays fundamental roles – location of mitotic cleavage plane – control of cell shape – directed transport of molecules Origin of germ line •In animals, germ line is set aside from soma in early development –only germ cells can undergo meiosis –somatic cells form body of organism •Asymmetric distribution of cytoplasmic particles (e.g., P granules of Caenorhabditis elegans) by cytoskeleton –cells receiving particles develop into germ line –particles anchored to actin in some organisms, to microtubules in others Drosophila anterior-posterior axis • Determined by gradients of BCD (product of bicoid) and HB-M (product of hunchback) – mRNA maternally deposited in egg – BCD mRNA tethered to “–” ends of microtubules via 3’ UTR – HB-M protein gradient depends on NOS protein • nos mRNA tethered to “+” end of microtubule via 3’ UTR • NOS protein gradient blocks translation of hb-m mRNA, resulting in HB-M gradient • Resulting opposite gradients of BCD and Nos determine axis Drosophila dorsal-ventral axis • Determined by gradient of transcription factor DL (encoded by dorsal) – gradient established by interaction of spz and Toll gene products deposited in oogenesis and released during embryogenesis – SPZ-TOLL complex triggers signal transduction pathway in cells that phosphorylates inactive DL • Phosphorylated DL migrates to nucleus, activating genes for ventral fates Positional information •Localization of mRNAs within cell establishes positional information in cases where developmental fields begin as a single cell •Formation of concentration gradients of extracellular diffusible molecules establishes positional information in multicellular developmental fields –works by signal transduction –diffusible molecules are known as morphogens Complex pattern: Drosophila •Successive interpretation of established, changing, and new gradients •Largely due to changes in transcription •Genes targeted by gradients of maternal A-P and D-V transcription factors are cardinal genes –respond to these factors at enhancers and silencers –similar genes in other animals Drosophila development (1) •Early syncitial development –zygotic nucleus divides 9 times with no cell division •some nuclei migrate to posterior pole to give rise to germ line –4 more mitotic divisions without cell division •Nuclei migrate to surface of egg cytoplasm –membrane forms around them (cellularization) –begin responding to positional information in A-P and D-V transcription factor gradients. Drosophila development (2) •At 10 hours, 14 segments –3 head –3 thoracic –8 abdominal •At 12 hours, organogenesis begins •At 15 hours, exoskeleton begins to form •At 24 hours, larva hatches Drosophila development (3) • Developmental fate determined through transcription-factor interactions • A-P cardinal genes = gap genes – Kruppel and knirps (mutants have gap in normal segmentation) – promoters have differential sensitivity to BCD and/or HB-M – establishes different developmental fields along embryo, roughly defining segments • Bifurcation of development: targets of gap gene encoded transcription factors – one branch to establish correct number of segments – one branch to assign proper identity to each segment Drosophila development (4) • Segment number – gap gene products activate pair-rule genes • several different pair-rule genes • expression produces repeating pattern of seven stripes, each offset – pair-rule products act combinatorially to regulate transcription of segment-polarity genes • expressed in offset pattern of 14 stripes • Segment identity – gap gene products target cluster of homeotic gene complexes • encode homeodomain transcription factors • mutations alter developmental fate of segment – e.g., Bithorax (posterior thorax and abdomen) and Antennapedia (head and anterior thorax) Pattern formation • Transcriptional response to gradients (asymmetrical distribution) of transcription factors • Memory of cell fate – intracellular and intercellular positive-feedback loops – e.g., homeodomain protein binds to enhancer elements of its own gene, ensuring continued transcription • Cell-cell interactions – inductive interaction commits groups of cells to same developmental fate – lateral inhibition results in neighboring cells assuming secondary fate Generalizations • Asymmetry of maternal gene products establishes positional information used for early development • Successive rounds of expression of genes encoding transcription factors establish axes and body part identity • Positive feedback loops maintain differentiated state • Components of one developmental pathway are also found in many others • Differences in types and concentrations of transcription factors result in different outputs Developmental parallels • Early animal development follows fundamentally similar pattern • Remarkable similarity among homeotic genes – one HOM-C cluster in insects – four HOX clusters in mammals • paralogous to insect cluster • expressed in segmental fashion in early development • Knockout and genome studies suggest animal development uses same regulatory pathways Development in plants • Plants have different organ systems than animals and plant cells can not migrate • Plants do not separate soma from germ-line until flower development • Plants too have hormones and signaling pathways • Arabidopsis thaliana is model system • Transcriptional regulators control fate of four whorls (layers) giving rise to flower – process similar action of homeotic genes in animal development Assignment: Concept map, Solved Problems 1 and 2, All Basic and Challenging Problems.